Studies on Chromatographic Fractioning by Cations Exchangers of a Bovine Hemoglobin Hydrolysate

Panainte, Alina Diana; Morariu, Ionela Daniela; Bibire, Nela; Vieriu, Madalina; Ţântaru, Gladiola; Luca, Alina Costina; Apostu, Mihai

doi:10.37358/rc.18.10.6626

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“…Anion and cation exchange chromatography is a common separation method used for tyrosinase purification [34][35][36]. A similar strategy was also used in our study.…”

Section: Separation By Ion Exchange Chromatographymentioning

confidence: 99%

A Biochemical Method for Tyrosine Determination in Phenylketonuria Using a Colorimetric Enzymatic Approach

Mocanu¹,

Bocec²,

Gradinaru³

et al. 2020

Rev. Chim.

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Phenylketonuria is a serious genetic disease caused by a deficiency of phenylalanine metabolism, an essential amino acid found in daily nutrition. This disorder is caused by the lack of a specific enzyme called phenylalanine hydroxylase which mediates the conversion of phenylalanine to tyrosine). Thus, after ingestion of phenyl alanine-rich proteins, the amino acid concentration increases considerably in the blood due to proteolysis. Genetic deffects of enzymes responsible for phenylalanine metabolic conversion were intensively studied. Among them the defect of gene encoding phenylalanine hydroxylase has a higher notoriety. This genetic defect is translated in an inactive enzyme constructs that impair the aminoacid hydroxylation. This physiological stage is also called hyperphenylalaninemia where slightly high levels of phenylalanine are noticed in the blood or urine. Consequently, the amino acid is converted to phenylpyruvic acid by transamination, the later displaying a particularly toxic effect against brain tissues. The aim of this study was to quantify tyrosine in the blood of patients suffering of phenylketonuria by an alternative enzymatic method. The tyrosinase used in this assay was extracted from commercial mushrooms (Agaricus bisporus) following the Haghbeen protocol with some modifications. Two chromatographic steps (molecular exclusion chromatography and ionic exchange chromatography) were used during the enzyme purification process. High purity samples were concentrated using ultrafiltration. The tyrosinase was screened by a classical enzymatic microplate assay having DOPA as a substrate. Finally the pure enzyme was used in order to quantify tyrosine from different standard solutions. The level of tyrosine from deproteinized serum samples was determined using a similar enzymatic strategy.

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“…Anion and cation exchange chromatography is a common separation method used for tyrosinase purification [34][35][36]. A similar strategy was also used in our study.…”

Section: Separation By Ion Exchange Chromatographymentioning

confidence: 99%